Genetic Variants Associated with Episodic Ataxia in Korea

Abstract

Episodic ataxia (EA) is a rare neurological condition characterized by recurrent spells of truncal ataxia and incoordination. Five genes (KCNA1, CACNA1A, CACNB4, SLC1A3, and UBR4) have been linked to EA. Despite extensive efforts to genetically diagnose EA, many patients remain still undiagnosed. Whole-exome sequencing was carried out in 39 Korean patients with EA to identify pathogenic mutations of the five known EA genes. We also evaluated 40 candidate genes that cause EA as a secondary phenotype or cerebellar ataxia. Eighteen patients (46%) revealed genetic information useful for establishing a molecular diagnosis of EA. In 11 patients, 16 pathogenic mutations were detected in three EA genes. These included nine mutations in CACNA1A, three in SLC1A3, and four in UBR4. Three patients had mutations in two genes, either CACNA1A and SLC1A3 or CACNA1A and UBR4, suggesting that SLC1A3 and UBR4 may act as genetic modifiers with synergic effects on the abnormal presynaptic activity caused by CACNA1A mutations. In seven patients with negative results for screening of EA genes, potential pathogenic mutations were identified in the candidate genes ATP1A2, SCN1A, TTBK2, TGM6, FGF14, and KCND3. This study demonstrates the genetic heterogeneity of Korean EA, and indicates that whole-exome sequencing may be useful for molecular genetic diagnosis of EA.

Figure 1

Pedigree of eight families with pathogenic mutations. Solid symbols (squares = males, circles = females) indicate clinically affected individuals; open symbols, unaffected individuals; and slashed symbols, deceased individuals. Probands are indicated by arrows.

Figure 2

Sequencing results and localization of the mutations in EA genes. (A) The Cav2.1 encoded by CACNA1A has four homologous domains (I-IV), each consisting of six transmembrane segments (S1-S6) and an additional pore loop located between S5 and S6. Three non-sense mutations (p.Arg1549TER of patient 16, p.Arg1669TER of patient 5 and p.Gln1986TER of patient 2) are predicted premature termination in the S4 of Domain IV or the C-terminal domain. A heterozygous insertion mutation is located in the intracellular linker connecting Domain II and III, and lead to a frameshift and premature termination (p.Gly1108Argfs40TER of patient 15). Two missense mutations (p.Gly677Glu of patient 1 and p.Tyr248Asn of patient 4) involve the pore region (S5-S6) of Domain II and I, respectively. The other missense mutation (p.Glu998Gln of patient 5) is located in the intracellular linker. The in-frame deletions without truncation of the protein (p.Glu1294DEL of patient 3 and p.Asp2202_Arg2205DEL of patient 15) are in the S2 of Domain III and the C-terminal domain, respectively. (B) The EAAT1 encoded by SLC1A3 is composed of eight alpha-helical transmembrane domains (TMDs) and re-entrant hairpin loops (HP) 1 and 2 flanking TMD7. The first sixth TMDs form a scaffold that surrounds a C-terminal core domain comprising HP1, TMD7, HP2, and TMD8. Two missense mutations (p.Ala329Thr of patient 3 and p.Thr318Ala of patient 23) are located in TMD6, while the other mutation (p.Val393Ile of patient 6) is located in TMD7, the critical binding site for glutamate and various coupled ions, Na⁺, H⁺ and K⁺. (C) The p600 protein encoded by UBR4 contains several identified functional domains including UBR box, microtubule (MT)-binding domains, and calmodulin (CaM)-binding domain. The two endoplasmic reticulum (ER)-binding regions are located near the center of the protein and within the MT-binding domain. The Ndel1-binding region overlaps with the MT-binding domain. Three missense mutations (p.Arg4111His of patient 5, p.Tyr4877Cys of patient 2, and p.Ala5042Val of patient 26) are located within the MT-binding domain, and one (p.Arg4111His) involves the CaM-binding domain interacting with calmodulin. The other one (p.Ala2581Val of patient 28) is situated in the functionally-unknown region.

Figure 3

Sequencing results and localization of the mutations in candidate genes. (A) The α2 subunit of the Na⁺/K⁺-ATPase encoded by ATP1A2, consists of 10 transmembrane segments (M1-M10) with a large cytoplasmic loop between M4 and M5. The missense mutation (p.Arg196Cys of patient 19) is located in the intracellular loop between M2 and M3. This region is associated with A- (actuator) domain responsible for the binding and hydrolysis of ATP. (B) The α1 subunit of the Na⁺ channel encoded by SCN1A, has four homologous domains (I-IV), each consisting of six transmembrane segments (S1-S6) and an additional pore loop located between S5 and S6. Two missense mutations (p.Ile1839Ser of patient 22 and p.Leu563Gln of patient 30) are located in the C-terminal domain and the intracellular linker connecting Domain I and II, respectively. (C) Missense mutations of genes associated spinocerebellar ataxia in four patients with episodic ataxia: TTBK2 (exon 15, c.3467 G > A, p.Arg1156Gln in patient 11), TGM6 (exon 10, c.1478 C > T, p.Pro493Leu in patient 12), FGF14 (exon 1, c.31 A > G, p.Thr11Ala in patient 32), KCND3 (exon 4, c.1291 C > T, p.Arg431Cys in patient 33).

Figure 4

Ictal and interictal cerebellar metabolism in patient 19 with ATP1A2 mutation. Compared with interictal PET (low panels), ictal images (upper panels) show significant hypometabolism in bilateral cerebellar hemispheres.